Topics in HIV MedicineÂ® - International AIDS Society-USA

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International AIDS Society–USA Topics in HIV Medicine Swift NK cell activation mediated through KIR molecules early in infection may positively impact viral control, not only for HIV, but also for other pathogens. Dendritic Cells Although DC-SIGN, a C-type lectin that is highly expressed in monocytederived dendritic cells (DCs), is believed to be a key molecule facilitating dendritic cell-mediated HIV infection of CD4+ T cells, the precise mechanisms underlying this function are not known. KewalRamani (Abstract 110) summarized research on DC-SIGN from his laboratory. Stable expression of DC-SIGN in the monocytic cell line THP-1 resulted in transmission of HIV-1 at efficiencies similar to those of primary immature DCs. In transwell experiments, cell contact between THP-1/DC-SIGN cells and CD4+ T lymphocytes was required for HIV-1 transmission. When other transformed cell lines (K562 and 3T3) stably transfected and expressing DC-SIGN at levels comparable to those of the THP- 1/DC-SIGN cells were examined, they were not able to transmit HIV-1, even though their ability to bind HIV-1 or intracellular adhesion molecule 3 (ICAM3) was not impaired. This finding led to a search for cell molecules required for DC-SIGN-mediated HIV transmission. As observed for immature DCs, HLA-DR and leukocyte function antigen 1 (LFA-1) were expressed on THP-1 cells permissible for HIV transmission, but these molecules were not expressed on K562 and 3T3 cells. However, expression of HLA class II molecules and LFA-1, singly or in combination, in the nonpermissive K562 and 3T3 cell lines did not reverse the defect in HIV transmission, suggesting that DC-SIGN expression is not sufficient for efficient HIV-1 transmission and that other cofactors are likely to play a role. Identification of these specific cofactors should provide valuable insights into DC-SIGN-mediated transmission of HIV. Studies using live-cell video microscopy to examine transfer of HIV from DCs to CD4+ T lymphocytes were presented by Hope (Abstract 113). Using fluorescent labeling of HIV by incorporation of a fusion protein of GFP with Vpr, investigators showed that in monocyte-derived DCs, HIV was recruited to sites of cell contact with T cells. At the same time, in the target cell, CD4 and CXCR4 were concentrated at the CD4+ T cell/DC interface. The localized concentration of HIV, CD4, and coreceptors (termed an “infectious synapse”) may play a critical role in facilitating infection of T cells by dendritic cells. Innate Immunity In addition to acting as a conduit for HIV transmission, DCs are the principal antigen-presenting cells in the body and essential for mounting an effective adaptive immune response. Loré and colleagues (Abstract 81) demonstrated that innate immune recognition mediated by binding of the microbial pattern recognition Toll-like receptors (TLRs) on DCs to their natural ligands resulted in maturation of DCs and enhancement of adaptive virus-specific immune responses in vitro. CD11c+ myeloid DCs expressing TLRs 3, 4, and 7 and CD123+ plasmacytoid DCs expressing TLRs 7 and 9 were activated on exposure to their respective ligands: CpG (TLR9 receptor), imidazoquinolones (TLR7), LPS (TLR4), and poly I:C (TLR3). TLR-ligand-mediated activation of plasmacytoid or myeloid DCs resulted in an enhanced capacity to generate antigen-specific T lymphocytes. The ability to modulate adaptive immunity makes TLR ligands a potentially useful immunologic adjuvant for HIV vaccines. Unutmaz (Abstract 112) outlined the role of natural killer T cells (NKT cells) in HIV infection. NKT cells are a distinct subset of human effector T lymphocytes that express an invariant T-cell receptor and recognize glycolipid antigens presented by the nonpolymorphic MHC class I molecule CD1d. Using flow cytometry to identify NKT cells in humans (Vα24+Vβ11+ or CD1d tetramer+Vβ11+), 20% to 90% of NKT cells were found to be CD4+ and to express CCR5. Autologous DCs pulsed with the glycolipid alpha-galactosylceramide were able to expand NKT cells, which secreted IL-4 and IL-5 and were highly susceptible to infection with R5-tropic HIV. HIV-infected subjects had marked declines in NKT cells, which may be mediated by direct infection. Decline in CD4+ NKT cells but not total NKT cells correlated inversely with plasma HIV-1 RNA level. Natural Hosts of SIV Infection An expanding number of nonhuman primates such as sooty mangabeys and African green monkeys serve as natural hosts for SIV infection. Despite lifelong infection and levels of viremia that can in some species equal or exceed those in HIV-infected subjects with AIDS, these hosts remain asymptomatic and do not develop AIDS. Increasing attention over the past several years has been devoted to trying to elucidate immunologic or virologic mechanisms associated with the lack of pathogenicity of primate lentiviruses in their natural hosts. Barry and colleagues (Abstract 120) proposed that a limited SIV-specific CD8+ T-cell response may play a causal role in the lack of immunodeficiency in sooty mangabeys naturally infected with SIVsm. Analysis of several markers of T-cell activation (eg, CD69, CD25, and HLA-DR) or cell proliferation (Ki67) revealed only minor increases in SIVsm-infected sooty mangabeys compared with those observed in uninfected mangabeys. Similar findings were observed even in the setting of acute SIVsm infection of sooty mangabeys, whereas acute infection of rhesus macaques with the same stock of SIVsm resulted in significant increases in T-cell activation, increased proliferation of CD8+ and CD4+ T cells, and progression to AIDS. Depletion of CD8+ T cells using a murine CD8-specific monoclonal antibody resulted in only 3-fold increases in plasma viremia, leading Barry and colleagues to conclude that a weak or absent SIV-specific CD8+ T-cell response may be an important factor in the lack of immunodeficiency in SIVinfected sooty mangabeys. A contrasting view was reported by Kaur (Abstract 121), who described the results of a study in which CD8+ T cells were depleted in SIV-infected sooty mangabeys using a chimeric mousehuman CD8-specific monoclonal antibody (cM-T807). This antibody has been widely used in nonhuman primate 80
Conference Highlights - HIV Pathogenesis and Vaccine Development Volume 11 Issue 3 May/June 2003 experiments and generally induces more prolonged and complete depletion of CD8+ T cells than has been obtained with murine monoclonal antibodies. Following a depletion of CD8+ T cells that often extended to 3 weeks, Kaur and colleagues observed up to 100-fold increases in plasma viremia in 5 of 6 animals studied, whereas no significant increases in viremia were observed in 4 animals that received a control chimeric monoclonal antibody. Further evidence for SIV-specific CD8+ T-cell responses was provided by the finding of significant ELISPOT responses to SIV proteins in most SIV-infected mangabeys. Reasons for the apparent discrepancy in findings between these groups was not immediately clear, but may relate to differences in the efficacy of CD8+ T-cell depletion induced by the 2 different monoclonal antibodies. Prophylactic AIDS Vaccines Neutralizing Antibody Responses After several years of being overshadowed by research on cell-mediated immune responses to HIV, research on neutralizing antibodies has recently enjoyed a renaissance prompted by improved information on the structural basis for neutralization, new assays for measuring neutralizing antibodies, and continued hope that structurally modified envelope immunogens will enhance the induction of broadly reactive neutralizing antibodies to HIV. Leading off a symposium devoted solely to neutralizing antibodies, Burton (Abstract 184) reviewed the evidence that supports the view that neutralizing antibodies represent the small fraction of envelope-specific antibodies that are able to bind the oligomeric envelope spike. According to this viewpoint, broadly reactive neutralizing antibodies are therefore the subset of antibodies that are able to bind to conserved sites on the oligomer. Burton also noted that although passive transfer of neutralizing antibodies in animal models has provided compelling proof-of-principle demonstration of the ability of neutralizing antibodies to protect against primate lentiviruses, efforts to elicit broadly neutralizing antibodies by vaccination have consistently met with failure. Describing one approach to better understand why it has been so difficult to induce broadly neutralizing antibodies by vaccination, Burton discussed detailed structural studies of 2 neutralizing monoclonal antibodies, b12 and 2G12. The first is a well-characterized neutralizing antibody that was initially isolated by a phage display library derived from an asymptomatic HIVinfected individual. Previous work had demonstrated that b12 recognized a conserved site of gp120 that overlaps with the CD4 binding site. A subsequent analysis of the crystal structure of b12 revealed a long protrusion at the tip of the antibody that inserts into the recessed CD4 binding site of gp120. In an effort to enhance production of antibodies with similar structure to b12, Burton and colleagues have tried to block production of antibodies to other common epitopes on the gp120 oligomer by hyperglycosylation of regions such as the CD4 binding site and the V3 loop. Immunization studies with the hyperglycosylated gp120 molecule are in process. A similar structural analysis has been carried out for the 2G12 antibody, which recognizes an epitope in the C4/V4 region of gp120 and has been previously shown to recognize a cluster of oligomannose glycans in this region of gp120. Crystal structure analysis of 2G12 revealed that instead of forming the typical Y shape of most immunoglobulin G (IgG) molecules, the hypervariable heavy chains (V H ) formed a tight cluster in which the 2 V H domains are directly juxtaposed, a phenomenon called domain swapping. The unusual structures of both b12 and 2G12 provide at least 1 reason for the difficulties in generating broadly neutralizing antibodies against HIV. Whether this understanding will ultimately facilitate elicitation of such antibodies by the use of modified immunogens will be a major challenge for research in this area for the next several years. After binding to cellular receptors, gp120 undergoes a series of conformational changes ultimately resulting in insertion of an oligomeric form of gp41 into the host cell membrane and subsequent fusion of the viral particle with the host cell membrane. Whether antibodies can be successfully generated to these intermediate forms of envelope, thereby blocking virus entry, has been controversial. Weiss (Abstract 185) reviewed efforts to generate antibodies able to block viral fusion. She identified 3 distinct stages at which fusion inhibitors may inhibit virus entry: prevention of conformational changes, blocking close opposition of the virus and host cell membrane, and prevention of fusion pore assembly. One of the intermediate steps of viral entry involves conformational changes in the ectodomain of gp41, in which 2 heptad repeats of gp41 are brought together to make up the prehairpin intermediate. This hairpin intermediate subsequently generates a 6-helix hairpin structure that catalyzes formation of the fusion pore. Peptides mimicking the gp41 heptad repeats are able to potently inhibit HIV replication, which suggests that antibodies might also be able to block this critical step in virus entry. Inherent challenges to this approach include steric hindrance (ie, whether antibodies can fit in the small gap between the virus particle and the host cell membrane) and whether sufficient time exists during viral fusion to allow the antibody to bind. Immunization with recombinant immunogens that mimic either the prehairpin or 6-helix bundle conformation of gp41 resulted in antibodies able to bind either early intermediates or late intermediates, respectively, and that were able to inhibit viral entry. These data provide an important proof-of-principle demonstration of the feasibility of inducing antibodies able to bind to these fusion intermediates. Generation of antibodies able to bind the chemokine coreceptor binding site of gp120 has been another leading approach for the induction of broadly neutralizing antibodies against HIV-1. Sodroski (Abstract 186) described recent efforts to define the structural characteristics of antibodies able to bind the chemokine receptor binding site. Two specific regions of gp120 make up the chemokine receptor binding site: the variable V3 loop and the beta19 strand, a more conserved region of the bridging sheet that is protected by the V3 loop. Because the length of IgG molecules (approximately 115 angstroms) is significantly longer than that of the space between gp120 and 81
Page 1 and 2: Topics in HIV Medicine ® Volume 11
Page 3 and 4: Table of Contents Volume 11 Issue 3
Page 5 and 6: Conference Highlights - Welcome Rem
Page 7 and 8: Conference Highlights - Basic Scien
Page 13 and 14: Conference Highlights - HIV Pathoge
Page 15: Conference Highlights - HIV Pathoge
Page 23 and 24: Conference Highlights - Complicatio
Page 29 and 30: Special Contribution - Drug Resista
Page 31 and 32: Special Contribution - Drug Resista
Page 33 and 34: Conference Highlights - Advances in
Page 65 and 66: Conference Highlights - Abstracts C
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Conference Highlights - Abstracts C
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Conference Highlights - Abstracts C
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